Hematopoietic stem cell transplantation (HSCT) has become the standard therapeutic approach for diverse malignant hematopoietic disorders, accounting with high rates of clinical success for chronic (80%) and acute myeloid leukemia (65%) due to long term hematopoietic engraftment and graft versus leukemia effects of the donor graft. However, post-HSCT subjects suffer from profound cellular immunodeficiency the first 100 days post-transplantation and reach full T and B cell reconstitution only after 1 year post-HSCT. Longer periods of cell immune deficiency are associated with subsequent increase of risk for opportunistic viral and fungal infections and relapse (up to 40%) (Seggewiss, R., and H. Einsele, 2010, “Immune reconstitution after allogeneic transplantation and expanding options for immunomodulation: an update.” Blood 115: 3861-3868; Roncarolo, M. G. et al., 2011, “Clinical tolerance in allogeneic hematopoietic stem cell transplantation.” Immunological reviews 241: 145-163; Mori, T., and J. Kato, 2010, “Cytomegalovirus infection/disease after hematopoietic stem cell transplantation” International journal of hematology 91: 588-595). Thus, strategies to accelerate the recovery of the lymphocyte pool with a broad repertoire of T cell and B cell responses and induce optimal viral immunity in post-HSCT transplanted subjects are required.
Novel advanced therapeutic approaches to induce immune reconstitution in immunodeficient hosts based on passive and active immunization have been developed over the past decade. Yet, suitable in vivo experimental models to address efficacy and biosafety of such therapies are still under development. In order to experimentally recapitulate human immune reconstitution after HSCT in vivo, CD34+ human hematopoietic stem cells (HSC) are transplanted into diverse immunodeficient mouse strains lacking the common interleukin-2 receptor gamma chain (IL2Rγ) (NOD-Rag1nullIL2Rγnull-NRG, NOD/LtSz-scid/IL2Rγnull-NSG, or NOD/SCID/IL2Rγnull-NOG) after sublethal total body irradiation (TBI), resulting in reconstitution of human hematopoietic lineages 8 to 10 weeks after CD34+ cell transfer (Ishikawa, F., et al., 2005, “Development of functional human blood and immune systems in NOD/SCID/IL2 receptor γ chainnull mice.” Blood 106: 1565-1573). Importantly, regardless the source of HSCs and the method of cell transplantation, humanized mice displayed suboptimal levels of T and B mature lymphocyte reconstitution, lack of antigen specific cellular and humoral responses and overall anergy (Lepus, C. M. et al., 2009, “Comparison of Human Fetal Liver, Umbilical Cord Blood, and Adult Blood Hematopoietic Stem Cell Engraftment in NOD-scid/γc−/−, Balb/c-Rag1−/−γc−/−, and C.B-17-scid/bg Immunodeficient Mice.” Human immunology 70: 790-802; Andre, M. C. et al, 2010, “Long-term human CD34+ stem cell-engrafted nonobese diabetic/SCID/IL-2R gamma(null) mice show impaired CD8+ T cell maintenance and a functional arrest of immature NK cells.” J Immunol 185: 2710-2720). Factors that determine the inefficient lymphatic development in humanized mice include the absence of human histocompatibility molecules, impaired thymic function and poor human cytokine environment.
Attempts to solve this problem included delivery of cytokines (O'Connell, R. M. et al., 2010, “Lentiviral Vector Delivery of Human Interleukin-7 (hIL-7) to Human Immune System (HIS) Mice Expands T Lymphocyte Populations.” PLoS One 5; Chen, Q. et al., 2009, “Expression of human cytokines dramatically improves reconstitution of specific human-blood lineage cells in humanized mice.” Proc Natl Acad Sci USA 106: 21783-21788), transplantation of fetal lymphatic tissue along with HPCs (Hu, Z., and Y. G. Yang, 2012, “Human lymphohematopoietic reconstitution and immune function in immunodeficient mice receiving cotransplantation of human thymic tissue and CD34+ cells” Cell Mol Immunol 9: 232-236; Biswas, S. et al., 2011, “Humoral immune responses in humanized BLT mice immunized with West Nile virus and HIV-1 envelope proteins are largely mediated via human CD5+ B cells.” Immunology 134: 419-433) and the use of transgenic strains constitutively expressing the major histocompatibility molecules (MHC) class I (Shultz, L. D. et al, 2010, “Generation of functional human T-cell subsets with HLA-restricted immune responses in HLA class I expressing NOD/SCID/IL2rγnull humanized mice.” Proc Natl Acad Sci USA 107: 13022-13027) and HLA class II (Danner, R. et al., 2011, “Expression of HLA Class II Molecules in Humanized NOD.Rag1KO.IL2RgcKO Mice Is Critical for Development and Function of Human T and B Cells” PLoS One 6) or critical hematopoietic cytokines (Willinger, T. et al., 2011, “Human IL-3/GM-CSF knock-in mice support human alveolar macrophage development and human immune responses in the lung” Proc Natl Acad Sci USA 108: 2390-2395) have been recently described. These strategies allowed a limited improvement in B and T cell responses against human viral challenges. Importantly, only few reports have described the presence of reconstituted lymphatic structures in HSC-transplanted mice (Singh, M. et al., 2012, “An Improved Protocol for Efficient Engraftment in NOD/LTSZ-SCIDIL-2RγNULL Mice Allows HIV Replication and Development of Anti-HIV Immune Responses” PLoS One 7; Marodon, G. et al., 2009, “High diversity of the immune repertoire in humanized NOD.SCID.gamma c−/− mice” European journal of immunology 39: 2136-2145; Sun, Z. et al., 2007, “Intrarectal transmission, systemic infection, and CD4+ T cell depletion in humanized mice infected with HIV-1” J Exp Med 204: 705-714). Although high quality CD34+HPC from human cord blood or fetal liver were used, thereby reaching high rates of human cell engraftment and reasonable levels of human lymphatic cells, lymph nodes (LN) were barely observed. These data suggest that lack of lymphatic organ regeneration could be playing an important role in the poor lymphoid cell reconstitution observed in humanized mouse models.
DCs play a central role in the induction of adaptive immune responses. Importantly, DC trigger the regeneration and remodeling of tertiary lymphatic structures and play a fundamental role in maintaining the function of LN during active immune responses.
Using lentivirus (LV)-mediated gene transfer, we have developed a method to generate highly viable and potent DCs for cancer immunotherapy (Pincha, M. et al., 2011, “Lentiviral vectors for induction of self-differentiation and conditional ablation of dendritic cells” Gene therapy 18: 750-764; Koya, R. C. et al., 2007 “Lentiviral vector-mediated autonomous differentiation of mouse bone marrow cells into immunologically potent dendritic cell vaccines” Molecular Therapy 15: 971-980″. LV-induced DC showed high levels of engraftment and potent capacity to stimulate antigen specific responses and protect against melanoma in vivo. Recently, we demonstrated that integrase-defective (ID) LV gene delivery of human granulocyte-macrophage colony stimulation factor (GM-CSF) and interferon (IFN)-α into human monocytes resulted in autonomous differentiated and highly viable dendritic cells (Daenthanasanmak, A. et al., 2012, “Integrase-defective lentiviral vectors encoding cytokines induce differentiation of human dendritic cells and stimulate multivalent immune responses in vitro and in vivo” Vaccine 30: 5118-5131).
Thus, the problem underlying the present invention can be viewed as the provision of means and methods which improve the regeneration of the immune system after transplantation of hematopoietic stem cells.
The problem is solved by the embodiments described in the claims and the description below.